Dataset Information

3-D Scaffold Platform for Optimized Non-viral Transfection of Multipotent Stem Cells.

ABSTRACT: Optimization of non-viral gene delivery from biomaterials is of critical importance, as several material parameters are known to influence non-viral transfection efficiency. A series of previous studies have achieved screening of gene delivery vectors on two dimensional (2D) substrates, which have direct relevance to cell culture applications. There is an additional need to create screening systems that are 3-dimensional (3D), and can thus be applied to emerging tissue engineering applications. Here, we report an enhanced throughput, 3D scaffold platform to screen for the influence of mineral coating properties on stem cell transfection. Mineral coatings with a range of physicochemical properties were formed on the scaffolds within a 96-well plate format, while maintaining an interconnected macroporous scaffold structure. A series of general gene delivery parameters, including plasmid amount, N/P ratio, and cell density, were efficiently screened in scaffolds using a luciferase-encoding plasmid as a reporter. In addition, human mesenchymal stem cell (hMSC) transfection with a plasmid encoding bone morphogenetic protein-2 (BMP-2) was successfully optimized by screening a library of mineral coatings, resulting in over 5-fold increases in BMP-2 production when compared to standard techniques. Notably, the majority of BMP-2 was incorporated into the mineral coating following secretion from the cells. The 3D mineral coated scaffold platform described here may accelerate gene delivery optimization and improve the predictability of the screening systems, which could facilitate translation of gene delivery to clinical applications.

SUBMITTER: Yu X

PROVIDER: S-EPMC4273581 | biostudies-literature | 2014 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

3-D Scaffold Platform for Optimized Non-viral Transfection of Multipotent Stem Cells.

Yu Xiaohua X Murphy W L WL

Journal of materials chemistry. B 20141201 46

Optimization of non-viral gene delivery from biomaterials is of critical importance, as several material parameters are known to influence non-viral transfection efficiency. A series of previous studies have achieved screening of gene delivery vectors on two dimensional (2D) substrates, which have direct relevance to cell culture applications. There is an additional need to create screening systems that are 3-dimensional (3D), and can thus be applied to emerging tissue engineering applications. ...[more]

PMID: 25541592

Similar Datasets

Project description:The pathogenesis of Rb1 gene inactivation indicates that gene therapy could be a promising treatment for retinoblastoma. An appropriate gene transfer system is the basis for successful gene therapy; however, little attention has been given to an effective gene transfer system for retinoblastoma therapy in previous studies. This study was designed to provide an optimized transgene system for WERI‑Rb1 cells (W-RBCs). Green fluorescent protein (GFP) was adopted as a reporter. Four classic viral vectors based on retroviruses, recombinant adeno-associated viruses (rAAV2, rAAV2/1), lentiviruses (LVs) and a novel non-viral vector X-treme HP reagent were adopted for W-RBC gene transfection. The efficacy and cytotoxicity were comprehensively compared among the different vectors through GFP expression and the trypan blue exclusion test. Furthermore, the serum and cell culture status were also optimized for better transfection. Cells transfected by rAAV2/1 expressed more GFP protein and exhibited less staining with trypan blue, compared to the rAAV2 counterpart. However, in comparison to the retroviral group, both the rAAV2/1 and LV groups had considerably less GFP+ cells. Interestingly, the X-treme HP presented a similar GFP transfection capacity to the retroviral vector, but with a much lower cytotoxicity. Furthermore, there were more GFP+ cells in a suspended condition than that in an adherent culture. Moreover, cells in a serum-positive system expressed more GFP, while cells in a serum-free system showed lower GFP expression and higher cytotoxicity. In conclusion, the retroviral vector and the X-treme HP are effective for W-RBC gene transfection, while the X-treme HP is more preferable due to its lower cytotoxicity. Moreover, the suspended cell culture system is superior to the adherent system, and the serum protects cell viability and facilitates the gene transfection of W-RBCs. This study presents an effective, convenient, and low toxic transfection system for gene delivery in W-RBCs and provides a promising system for further gene therapy of retinoblastoma.

Dataset Information

3-D Scaffold Platform for Optimized Non-viral Transfection of Multipotent Stem Cells.

Publications

3-D Scaffold Platform for Optimized Non-viral Transfection of Multipotent Stem Cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets